We are investigating the unique chromosomal metabolic events associated with meiosis and the repair of chromosomes following exposure to DNA damaging agents. Many of the genes necessary for the repair of DNA double strand breaks (DSB) are required for the successful completion of the meiotic cycle. Two proteins/enzyme have been identified that appear to play a role in meiosis and/or repair; a Mg(2+) dependent nuclease (RhoNUC, previously referred to as yNucR, and a protein that is able to carry out a strand exchange reaction (SEP). Both of these protein activities appears to under the control of the RAD52 gene, a gene required for the repair of radiation-induced DSBs and the completion of meiosis. To investigate the role of RAD52, we have created strains containing a chromosomal resident deletion of the RAD52 and a RAD52 gene on a plasmid under the control of the yeast GALl promoter. This allows us to tightly regulate the expression of the RAD52 gene. We have found that for the repair of DSBs the presence of RAD52 protein is required prior to treatment with ionizing radiation. Interestingly, the expression of the RhoNUC protein does not require a functional RAD52 gene. This suggests the control of RhoNUCH by RAD52 is not at the transcriptional or translational level. In order to identify other proteins which may interact with the RAD52 protein, we have begun screening a "protein interaction" library. This system will allow us to identify other proteins which physically interact with RAD52. The possible role of RAD52 during DNA replication is also being investigated. Using a unique system whereby we measure the frequency of "excision" of a bacterial transposon Tn5 placed into the yeast LYS2 gene, we have found the mutations in the genes encoding DNA polymerase I or III lead to high levels of Tn5 "excision". No increase is observed in DNA polymerase II mutants. The elevated levels of "excision" seen in polIII or polI strains is reduced when a null mutant of RAD52 is introduced into these strains. This suggests an interaction between the enzymes proposed to be responsible for DNA lagging strand synthesis (POLIII and POLI) and RAD52. Currently we are investigating the nature of this interaction.